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Protease-Activated Receptor Isoform Expression in Pregnant and Nonpregnant Rat Myometrial Tissue

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Abstract

Objective

Three protease-activated receptor (PAR1, 3, and 4) isoforms have been shown to be responsible for the cellular effects of thrombin; another PAR isoform (PAR2) is responsible for the cellular effects of trypsin. The present studies sought to test the hypothesis that one (or more) of these PAR isoforms is expressed in myometrial tissue, thereby accounting for the uterotonic effects of these novel agonists.

Methods

The rat PAR3 and 4 isoforms were cloned from a rat spleen cDNA library. PAR isoform mRNA expression was determined by using reverse-transcriptase polymerase chain reactions (PCR) in Sprague-Dawley rats. Confirmation of the identity of the amplified mRNA was done by sequence analysis. Relative quantification of the PAR1 and PAR2 isoforms was performed using a real-time quantitative reverse transcriptase PCR (RT-PCR) technique. PAR protein expression was confirmed by Western blots using polyclonal antibodies.

Results

The rat PAR3 and 4 homologues showed significant sequence homology to the mouse and human amino acid and nucleotide sequences. The RT-PCR studies confirmed PAR1-4 expression in myometrium from rats in estrus. PAR3 was expressed in uterus, spleen, kidney, liver, lung, brain, and heart. PAR4 was expressed in uterus, spleen, and lung. Messenger RNAfor the PAR1 and 2 isoforms was expressed during the second half of gestation in myometrium from timed-pregnant rats. In contrast, mRNA for the PAR3 and 4 isoforms was not detected in gestational myometrium. PAR protein expression appeared to match tissue mRNA expression patterns.

Conclusion

These RT-PCR studies confirmed ubiquitous expression of the PAR1 and PAR2 isoforms in myometrium and other rat tissues; in contrast, the PAR3 and PAR4 isoforms are expressed in a tissue-specific and gestationally related pattern.

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Authors and Affiliations

  1. Department of Obstetrics and Gynecology, University of Vermont, Given Building C258, 89 Beaumont Drive, Burlington, Vermont, 05405, USA

    Edward K. Chien MD, Leigh Sweet, Mark Phillippe MD, Sarah Marietti MD, Terrence T. Kim MD, David A. Wolff MS, Leandra Thomas MS & Eric Bieber MD

  2. University of Arizona College of Medicine, Tucson, Arizona, USA

    Edward K. Chien MD, Leigh Sweet, Mark Phillippe MD, Sarah Marietti MD, Terrence T. Kim MD, David A. Wolff MS, Leandra Thomas MS & Eric Bieber MD

  3. Department of Obstetrics and Gynecology, University of Chicago, Chicago, Illinois, USA

    Edward K. Chien MD, Leigh Sweet, Mark Phillippe MD, Sarah Marietti MD, Terrence T. Kim MD, David A. Wolff MS, Leandra Thomas MS & Eric Bieber MD

  4. Department of Obstetrics and Gynecology, Northwestern University, Chicago, Illinois, USA

    Edward K. Chien MD, Leigh Sweet, Mark Phillippe MD, Sarah Marietti MD, Terrence T. Kim MD, David A. Wolff MS, Leandra Thomas MS & Eric Bieber MD

  5. Department of Obstetrics and Gynecology, Geisinger Health System, Danville, Pennsylvania, USA

    Edward K. Chien MD, Leigh Sweet, Mark Phillippe MD, Sarah Marietti MD, Terrence T. Kim MD, David A. Wolff MS, Leandra Thomas MS & Eric Bieber MD

Authors
  1. Edward K. Chien MD
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  2. Leigh Sweet
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  3. Mark Phillippe MD
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  8. Eric Bieber MD
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Corresponding author

Correspondence to Edward K. Chien MD.

Additional information

This research was funded by grants from the National Institute of Child Health and Human Development: HD01232-01 (EKC), HL07479 (SM), HD32449 (MP), and HD28506 (MP).

We thank Graeme Bell for the use of the ABI7700.

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Chien, E.K., Sweet, L., Phillippe, M. et al. Protease-Activated Receptor Isoform Expression in Pregnant and Nonpregnant Rat Myometrial Tissue. Reprod. Sci. 10, 460–468 (2003). https://doi.org/10.1016/S1071-55760300148-5

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